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Figure 3.
(a) Superposition of phosphopeptide binding pockets of
PBD^PL, PBD^PP, PBD^S+G and PBD^S. Gray, PBD; green, PLHSpT;
yellow, PLHSpT-associated glycerol molecule; cyan, PPHSpT;
magenta, glycerol molecule (two half-occupancy conformations at
Ser–1 position) of PBD^S+G; black, two sulfate anions of
PBD^S+G and PBD^S (red, oxygen atoms). Differences in exact
positions of sulfate and phosphate groups could result from the
fact that sulfate is a free anion, whereas phosphate is
covalently linked to the phosphopeptide. (b) PBD residues
involved in binding of PLHSpT are labeled and shown in cyan. All
water molecules that form an interface between the
phosphopeptide and PBD are drawn in red mesh. (c) Superposition
of PLHSpT (green), PPHSpT (cyan), MQSpTPL (magenta) and PMQSpTPL
(gray). (d,e) Mixture of HeLa lysates expressing kinase-inactive
Flag-PLK1(K82M), Flag-PLK2(K108M) or Flag-PLK3(K52R) was
subjected to pull-down assays as in Figure 2a, with the
indicated 5-mer wild-type (PLHSpT) and mutants cross-linked to
beads. The nonphosphorylated Thr78 peptide PLHST was used as a
control. Numbers above the blot indicate relative efficiency of
PLK2 precipitation; numbers below denote relative efficiency of
PLK1 precipitation. (f) Nature of interactions between
SpT-containing peptides and PLK1 PBD. Alignment of minimal p-T78
peptides (PLHST and LHSTA) and synthetic optimal peptides
(PMQSTPL and MQSTPL) are shown. See text for details.
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